Mycobacterium tuberculosis causes the contagious disease tuberculosis and has infected up to one-third of the global population and remains one of the leading causes of fatal infections. The disease is responsible for an estimated two million deaths annually. Drug resistant strains of M tuberculosis have been steadily increasing in frequency, posing an additional and more serious threat to public health. The bacterium enters and resides in its host cell macrophage by subverting phagosomal processing, such that following engulfment the resulting phagosome compartment remains at an early endosomal stage rather than maturing into a bactericidal phagolysosome. Several mechanisms have been proposed for this critical aspect of pathogenesis, and it seems certain that there are multiple means by which M tuberculosis prevents phagosomal maturation. As a result, M. tuberculosis remains a widespread and devastating human pathogen.
There is evidence that mycobacterial-specific cell surface lipids play a role in the arrest of phagosome maturation, evidence for other effectors is less definitive. Deretic et al., Cell. Microbiol. 8:719 (2006). Different genetic screens indicate roles for non-overlapping sets of genes. One screen focused on the primary effects early in the infection process and highlighted a role for the product of a five-gene operon nominally involved in isoprenoid biosynthesis, with mutations in the two unique/non-redundant genes (Rv3377c and Rv3378c) leading to a significantly decreased ability to prevent phagosomal maturation. Pethe et al., Proc. Natl. Acad. Sci., 101(37):13642-13647 (2004).
Phagosomes containing wild-type M tuberculosis demonstrate an inability to acidify below pH 6.2. However, those containing the corresponding mutant mycobacteria acidified to pH 5.7, resulting in more than a three-fold increase in proton concentration causing a significant reduction in bacterial proliferation in macrophage cell culture. The fact that the mutants were among those with the most extreme phenotype, indicated that the product of the operon is required for M tuberculosis to attain full virulence and plays a role in the initial stages of M tuberculosis entry into macrophages. The enzyme encoded by the Rv3377c gene acts as a diterpene cyclase producing bicyclic halimadienyl diphosphate from the acyclic primary metabolite geranygeranyl diphosphate (GGPP) via a protonation-initiated (i.e., class II) cyclization mechanism. Nakano et al., Chem Comm, 2005:1016-1018 (2005).
The pathogenesis of various pathogens, such as M. tuberculosis, has led to research in the area of natural products including labdane-related diterpenoids. Labdane diterpenoids comprise a large group of approximately 7,000 known natural products defined as minimally containing the fused bicyclic hydrocarbon structure found in the labdane family of diterpenoids. The characteristic core structure results from the unusual biosynthetic origins of the compounds, uniquely initiated by a sequential pair of terpene synthase catalyzed reactions. Although a number of labdane diterpenoids exhibit medically relevant effects, including antibiotic, anti-inflammatory and anti-cancer activity, there remains a need for additional research in the area of novel labdane diterpenoids and the biological activity of diterpenoids for a variety of medically-relevant purposes, including the identification of virulence factors of known pathogens.
Therefore, it is an object of the present invention, to identify, isolate and characterize the diterpene virulence factor of M tuberculosis. 
It is a further object of the present invention to develop compounds for use as immune modulators for M tuberculosis pathogens as well as immune system disorders, such as autoimmune diseases, allergies and other immune system conditions.
It is an additional object of the present invention to develop pharmaceutical compositions comprising the diterpene isotuberculosinol.
It is an additional object of the present invention to develop drug assays for compounds capable of inhibiting the production of the diterpene isotuberculosinol and/or terpene cyclases.
It is an additional object of the present invention to characterize a terpene cyclase to from M. tuberculosis capable of producing the diterpene virulence factor isotuberculosinol.
It is an additional object of the present invention to develop assay methods for the diterpene isotuberculosinol.
These and other objects of the invention will become more readily apparent from the following detailed description, examples and appended claims.